There is a demand for a rapid, simple, cost-effective technique for screening air, water and blood samples to identify various components therein. Screening can involve detection of harmful chemicals, bacteria and viruses. For example, there is a need for early medical diagnostics, genomics assays, proteomics analyses, drug discovery screening, and detection of biological and chemical warfare agents for homeland security and defense.
Screening can also be used to identify the presence or absence of medical diseases and infectious pathogens. Regarding blood, the use of inexpensive screening analyses can allow the rapid detection and improved treatments of many illnesses. Rapid and effective medical screening tests can also reduce the cost of health care by preventing unnecessary and costly reactive medical treatment.
A critical factor in many diagnostics is the rapid, selective, and sensitive detection of biochemical substances, such as proteins, metabolites, nucleic acids, biological species or living systems, such as bacteria, virus or related components at ultra-trace levels in samples provided. In the case of medical diagnostic applications, biological samples can include tissues, blood and other bodily fluids. To achieve the required level of sensitivity and specificity in detection, it is often necessary to use a biosensor that is capable of identifying and differentiating between a large number of biochemical constituents in complex samples.
Living systems possess exquisite recognition elements, such as antibodies, enzymes and genes, often referred to as bioreceptors, which allow specific identification and detection of complex chemical and biological species. Biosensors exploit this powerful molecular recognition capability of bioreceptors. Due to the high level of specificity of the DNA hybridization process, there is an increasing interest in the development of DNA bioreceptor-based analytical systems. Applications for these systems include infectious disease identification, medical diagnostics and therapy, biotechnology and environmental bioremediation.
There has been recent research and development relating to biosensors. One type of biosensor device, often referred to as a “biochip,” applies spectroscopy using a semiconductor-based detection system and biotechnology-based probes. Bioprobes have been receiving increasing interest as of late. These probes have generally included luminescence labels, such as fluorescent or chemiluminescent labels for gene detection. Although sensitivities achieved by luminescence techniques are generally adequate for certain applications, alternative techniques with improved spectral selectivities are desirable to overcome the limited spectral specificity generally provided by luminescent labels.
Spectroscopy is an analytical technique concerned with the measurement of the interaction of radiant energy with matter and with the interpretation of the interaction both at the fundamental level and for practical analysis. Interpretation of the spectra produced by various spectroscopic instrumentation has been used to provide fundamental information on atomic and molecular energy levels, the distribution of species within those levels, the nature of processes involving change from one level to another, molecular geometries, chemical bonding, and interaction of molecules in solution. Comparisons of spectra have provided a basis for the determination of qualitative chemical composition and chemical structure, and for quantitative chemical analysis.
Vibrational spectroscopy is a useful technique for characterizing molecules and for determining their chemical structure. The vibrational spectrum of a molecule, based on the molecular structure of that molecule, is a series of sharp lines which constitutes a unique fingerprint of that specific molecular structure. If the vibrational spectrum is to be measured by an optical absorption process, optical fibers from a source is delivered to a sample, and after passage through the sample, an optical signal generated by the exciting optical energy is collected. This collected light is directed to a monochromator equipped with a photodetector for analyzing its wavelength and/or intensity.
One particular spectroscopic technique, known as Raman spectroscopy, utilizes the Raman effect, which is a phenomenon observed in the scattering of light as it passes through a material medium, whereby the light experiences a change in frequency and a random alteration in phase. When light is scattered from a molecule, most photons are elastically scattered. The scattered photons have the same energy (frequency) and, therefore, wavelength, as the incident photons. However, a small fraction of light (approximately 1 in 107 photons) is scattered at optical frequencies different from, and usually lower than, the frequency of the incident photons. The process leading to this inelastic scatter is termed the Raman effect. Raman scattering can occur with a change in vibrational, rotational or electronic energy of a molecule.
The difference in energy between the incident photon and the Raman scattered photon is equal to the energy of a vibration of the scattering molecule. A plot of intensity of scattered light versus energy difference is a Raman spectrum. The wavelengths present in the scattered optical energy are characteristic of the structure of the molecule, and the intensity of this optical energy is dependent on the concentration of these molecules.
Numerically, the energy difference between the initial and final vibrational levels, v, or Raman shift in wavenumbers (cm−1), is calculated through equation 1 below:v=(1/λincident)−(1/λscattered)  (1 )
Where λincident and λscattered are wavelengths (in cm) of the incident and Raman scattered photons, respectively. The vibrational energy is ultimately dissipated as heat. Because of the low intensity of Raman scattering, heat dissipation does not cause a measurable temperature rise in the material.
Raman spectroscopy is complementary to fluorescence, and has been used as an analytical tool for certain applications due to its excellent specificity for chemical group identification. However, low sensitivity historically has limited its applications.
Recently, the Raman technique has been rejuvenated following the discovery of a Raman enhancement of up to 106 to 1010 for molecules adsorbed on microstructures of metal surfaces. The technique associated with this phenomenon is known as surface-enhanced Raman scattering (SERS) spectroscopy. The enhancement is due to a microstructured metal surface scattering process which increases the intrinsically weak normal Raman scattering (NRS) due to a combination of several electromagnetic and chemical effects between the molecule adsorbed on the metal surface and the metal surface itself.
The enhancement is primarily due to plasmon excitation at the metal surface. Thus, the effect is generally limited to Cu, Ag and Au, and to a few other metals for which surface plasmons are excited by visible radiation. Although chemisorption is not essential, when it does occur there may be further enhancement of the Raman signal, since the formation of new chemical bonds and the consequent perturbation of adsorbate electronic energy levels can lead to a surface-induced resonance effect. The combination of surface- and resonance-enhancement (SERS) can occur when adsorbates have intense electronic absorption bands in the same spectral region as the metal surface plasmon resonance, yielding an overall enhancement as large as 1010 to 1012.
Raman spectroscopy has become an important analytical technique for chemical and biological analysis due to the wealth of information on molecular structures, surface processes, and interface reactions that can be extracted from experimental data. The Raman technique has been used with gene probe biosensors. U.S. Pat. No. 5,814,516 ('516 patent) to the same Inventor as the instant invention entitled “Surface enhanced Raman gene probe and methods thereof” discloses a gene probe biosensor comprising a support means, a SERS gene probe having at least one oligonucleotide strand having at least one SERS label, and a SERS active substrate disposed on the support means. The support means has at least one SERS gene probe adsorbed thereon. Biotargets such as bacterial and viral DNA, RNA and PNA are detected using a SERS gene probe via hybridization to oligonucleotide strands complementary to the SERS gene probe. The '516 patent does not disclose or suggest operatively connecting a Raman gene probe with an integrated circuit detection system to produce a biochip capable of SERS detection.